Selenophosphate (SeP), the energy-rich Se compound required for synthesis of many selenoenzymes is formed by selenophosphate synthetase (SPS) from ATP and an inorganic form of Se. The purified enzyme contains a bound UV absorbing chromophore characterized by Dr. Matt Wolfe. The participation of specialized selenium delivery proteins to furnish Se specifically to SPS serves to maintain intracellular levels of selenium below toxic levels. Certain classes of delivery proteins that can utilize selenocysteine as substrate, selenocysteine lyases, include three NifS related proteins from E. coli and one from Methanococcus vannielii, an anaerobic organism that is particularly rich in selenoenzymes and factors required for selenoprotein biosynthesis. A selenium-binding protein that reacts with inorganic selenium was isolated from M. vannielii and the N-terminal sequence determined. The gene encoding this protein was isolated, cloned, and expressed in E. coli by Dr. William Self. The expression protein is currently being studied by Dr. Kemberly Patteson. The expressed protein binds selenite. Antibodies to this protein allow its ready isolation from M. vannielii crude extracts. To date no stabilized forms or adducts of the highly oxygen-labile selenophosphate have been found either in mammalian or bacterial systems. Either this general selenium donor compound must be synthesized in situ wherever needed or some transportable form should exist. Based on properties of a thiophosphate-menadione addition product that we prepared previously, a corresponding derivative of selenophosphate might exist. The gene encoding tRNA-2-selenouridine synthase was expressed in E. coli. The selenium donor for 2-selenouridine formation is selenophosphate. The formation of 2-selenouridine involves replacement of sulfur in 2-thiouridine with Se. The enzyme and the reaction mechanism are being investigated by Dr. Matt Wolfe. A group of selenium-dependent molybdopterin hydroxylases present in some anaerobic bacteria contain an unidentified cofactor form of selenium. A purine hydroxylase discovered and studied by Dr. William Self is an excellent system for study of this selenium cofactor, particularly as regards its mode of synthesis and mechanism of addition to these enzymes. Selenophosphate as possible donor of Se for the hydroxylase cofactor is being investigated.